Human/Mouse/Rat Phospho-ERK1 (T202/Y204)/ERK2 (T185/Y187) Antibody

Detection of Mouse, Rat, and Human Phospho-ERK1 (T202/Y204) and ERK2 (T185/Y187) by Western Blot. Western blot shows lysates of NIH-3T3 mouse embryonic fibroblast cell line untreated (-) or treated (+) with 100 ng/mL Rabbit Anti-Human PDGF (Catalog # 120-HD) and PC-12 rat adrenal pheochromocytoma cell line untreated or treated with 100 ng/mL Recombinant Rat beta -NGF (Catalog # 556-NG) and HeLa human cervical epithelial carcinoma cell line untreated or treated with 200 nM PMA. PVDF membrane was probed with 0.5 µg/mL of Human/Mouse/Rat Phospho-ERK1 (T202/Y204)/ERK2 (T185/Y187) Monoclonal Antibody (MAB1018), followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). Specific bands were detected for Phospho-ERK1 (T202/Y204)/ ERK2 (T185/Y187) at approximately 40-45 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 3.

Phospho-ERK1 (T202/Y204)/ERK2 (T185/Y187) in HeLa Human Cell Line. ERK1/ERK2 phosphorylated at T202/Y204 (ERK1) and T185/Y187 (ERK2) was detected in immersion fixed Hela human cervical epithelial carcinoma cells, unstimulated (lower panel) and stimulated (upper panel) with PMA using Rabbit Anti-Human/Mouse/Rat Phospho-ERK1/ERK2 (ERK1 T202/Y204, ERK2 T185/Y187) Monoclonal Antibody (Catalog # MAB1018) at 10 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Rabbit IgG Secondary Antibody (red; Catalog # NL004) and counterstained with DAPI (blue). Specific staining was localized to cytoplasm. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

Detection of Phospho-ERK1/ERK2 in PMA-treated Jurkat Human Cell Line by Flow Cytometry. Jurkat human acute T cell leukemia cell line were unstimulated (light orange open histogram) or treated with 50 ng/mL PMA for 10 minutes (dark orange filled histogram), then stained with Rabbit Anti-Human/Mouse/Rat Phospho-ERK1/ERK2 (ERK1 T202/Y204, ERK2 T185/Y187) Monoclonal Antibody (Catalog # MAB1018) or control antibody (Catalog # AB-105-C, blue open histogram), followed by Fluorescein-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # F0112). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with ice-cold methanol.

Detection of Human Phospho-ERK1 (T202/Y204)/ERK2 (T185/Y187) by Simple Western^TM. Simple Western lane view shows lysates of Jurkat human acute T cell leukemia cell line untreated (-) or treated (+) with 200 nM PMA and Ionomycin for 20 minutes, loaded at 0.2 mg/mL. A specific band was detected for Phospho-ERK1 (T202/Y204)/ERK2 (T185/Y187) at approximately 42 kDa (as indicated) using 5 µg/mL of Rabbit Anti-Human/Mouse/Rat Phospho-ERK1/ERK2 (ERK1 T202/Y204, ERK2 T185/Y187) Monoclonal Antibody (Catalog # MAB1018). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Detection of Human ERK1/ERK2 by Flow Cytometry Differentiation Characteristics of Individual Stem Cells.(A) Flow cytometric analysis of undifferentiated stem cell markers in the clonal NTERA2.Tom cell line. Black line depicts P3X negative control and green depicts antigen specific expression (OCT4, SOX2, NANOG and SSEA4). (B) Graph depicting the percentage of TUJ1 positive neurons within differentiated colonies derived from individual RA treated NTERA2.Tom stem cells. (C) Images of differentiated NTERA2.Tom colonies derived from individual NTERA2.Tom cells on a background of wildtype NTERA2 cells. Cell were exposed to RA for 21 days (Green - beta III tubulin, Blue - Hoechst) White arrows depict beta III tubulin positive NTERA2.Tom neurons. Scale bar represents 50 µM. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/20531938), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human ERK1/ERK2 by Western Blot p38 inhibitor SB203580 improves the up-regulation of MMP-1 by DDC through stimulating ERK1/2Co-cultures were treated with or without 100 μM DDC for 1 h in the presence or absence of p38 inhibitor (SB203580, 10 μM). Phosphorylation of ERK1/2, p38 and Akt in LX-2 cells of co-cultures were determined by Western blotting analysis. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/23577625), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human ERK1/ERK2 by Western Blot Phospho-proteome profiling results of LX-2 cells co-cultured with C3A–CYP2E1 cells with or without DDC treatment and detection of the effect of DDC on intracellular kinases in LX-2 cells of co-cultures200 μg of total cell lysates from LX-2 cells co-cultured with C3A-2E1 cells with or without 100 μM DDC for 1 h were incubated with membranes of the human phospho-MAPK Array Kit according to the manufacturer's instructions. Phospho MAPK Array data were developed on X-ray films following exposure to chemiluminescent reagents. 20 μg aliquots of total cell lysates from LX-2 cells were subjected to Western blotting analysis. (A) Template showing the location of MAPK antibodies spotted onto the human phospho-MAPK Array Kit. (B) The activation status of ERK1/2 in LX-2 cells co-cultured with C3A-2E1 cells after DDC treatment. (C) The activation status of p38 in LX-2 cells co-cultured with C3A-2E1 cells after DDC treatment. (D) The activation status of Akt in LX-2 cells co-cultured with C3A-2E1 cells after DDC treatment. (E) The activation status of ERK1/2, p38 and Akt in LX-2 cells co-cultured with C3A cells after DDC treatment. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/23577625), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human ERK1/ERK2 by Western Blot DDC up-regulates MMP-1 through ERK1/2 and Akt activation(A) Co-cultures were treated with 100 μM DDC for the indicated time. Phosphorylation of ERK1/2, p38 and Akt were determined by Western blotting analysis. The corresponding non-phosphorylated ERK1/2, p38, Akt and beta -actin were used for protein loading control. (B) Co-cultures were treated with or without 100 μM DDC for 24 h in the presence or absence of ERK1/2 inhibitor (U0126, 10 μM). MMP-1 protein levels were analysed by Western blotting. (C) Co-cultures were treated with or without 100 μM DDC for 24 h in the presence or absence of p38 inhibitor (SB203580, 10 μM). MMP-1 protein levels were analysed by Western blotting. (D) Co-cultures were treated with or without 100 μM DDC for 24 h in the presence or absence of Akt inhibitor (T3830, 50 μM). MMP-1 protein levels were analysed by Western blotting. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/23577625), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human ERK1/ERK2 by Western Blot Phospho-proteome profiling results of LX-2 cells co-cultured with C3A–CYP2E1 cells with or without DDC treatment and detection of the effect of DDC on intracellular kinases in LX-2 cells of co-cultures200 μg of total cell lysates from LX-2 cells co-cultured with C3A-2E1 cells with or without 100 μM DDC for 1 h were incubated with membranes of the human phospho-MAPK Array Kit according to the manufacturer's instructions. Phospho MAPK Array data were developed on X-ray films following exposure to chemiluminescent reagents. 20 μg aliquots of total cell lysates from LX-2 cells were subjected to Western blotting analysis. (A) Template showing the location of MAPK antibodies spotted onto the human phospho-MAPK Array Kit. (B) The activation status of ERK1/2 in LX-2 cells co-cultured with C3A-2E1 cells after DDC treatment. (C) The activation status of p38 in LX-2 cells co-cultured with C3A-2E1 cells after DDC treatment. (D) The activation status of Akt in LX-2 cells co-cultured with C3A-2E1 cells after DDC treatment. (E) The activation status of ERK1/2, p38 and Akt in LX-2 cells co-cultured with C3A cells after DDC treatment. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/23577625), licensed under a CC-BY license. Not internally tested by R&D Systems.